The major objective of our research program is to determine regulatory roles of key enzymes and proteins involved in the biosynthesis, utilization and transport of cholesterol. Specifically, we are focusing our studies on three regulatory enzymes in cholesterol metabolism. In the area of cholesterol biosynthesis, we are studying HMG-CoA reductase and squalene synthetase. HMG-CoA reductase is the major rate-limiting step in cholesterol formation, while squalene synthetase is the committed step in sterol biosynthesis. In cholesterol utilization we are studying acyl-CoA cholesterol acyl transferase (ACAT), the enzyme which forms cholesterol esters from unesterified cholesterol and fatty acyl-CoA derivatives of fatty acids. Also being investigated is sterol carrier protein-2 (SCP-2), a protein which participates in cholesterol biosynthesis, utilization and intracellular transfer. Using techniques of site-directed mutagenesis, we will examine the role of SCP-2 structure in relation to its functional activity. It is known that there is a direct and exponential relationship between plasma cholesterol concentration and the risk of death due to coronary heart disease. Therefore, understanding the molecular mechanisms involved in regulating cholesterol metabolism is of key importance.